We propose to use electron spin resonance (ESR) and spin label techniques to probe the surface coat and lipid of the sperm plasma membrane to determine how membrane fluidity and permeability are modified during in vivo capacitation and various in vitro capacitating treatments. Water soluble spin probes and an appropriate broadening agent will be used to determine changes in volume and viscosity of the aqueous compartments, as well as, membrane porosity. The viscosity and physical state of membrane lipid will be probed using phospholipid, steroid and fatty acid spin labels. The surface coating of the plasma membrane will be examined with sialic acid and protein labeling spin probes. We will determine the relative amounts of sialic acid, protein and lipids that are removed during capacitation and whether there changes can be reversed by decapacitation. For sperm capacitated in vitro, we will determine if the plasma membrane is essential to initiate the increased respiration associated with capacitation or whether this response can be brought about directly in membrane-free or fractionated sperm by factors in the medium. If the membrane appears essential to the response we will isolate the membranes from noncapacitated sperm, subject them to in vivo capacitated membranes relative to controls. Parallel studies will be run for intact sperm and the magnitude of difference for adenylate cyclase levels between capacitated and control will be compared to those observed for the isolated membranes. We also will determine the biophysical effects of albumin and catecholamines on the sperm plasma membrane. We will investigate whether the apparent reduction in water volume observed by ESR for capacitated sperm and sperm from frequently ejaculated rabbits is associated with changes in membrane permeability to the spin probe induced by these treatments. These studies should improve our understanding of biophysical changes in sperm compartments and membrane integrity associated with capacitation. This information should be helpful in designing and testing contraceptive measures directed at the process of sperm capacitation and in correcting problems of infertility.